1. Field of the Invention
The present invention relates to a technique using a sample image obtained by scanning a sample with light via an optical system of a confocal scanning microscope, and by using light reflected from or passing through the sample, and more particularly, to a technique for relating a stereoscopic partial region in the sample to the sample image thus obtained.
2. Description of the Related Art
A confocal scanning microscope scans a sample with a spotlight source, converts only light, which passes through a pinhole among light beams reflected from and passing through the sample, into an electric signal with a photodetector, and obtains three-dimensional information of the sample from this electric signal.
Such a confocal scanning microscope suppresses light scattered from other than a measurement point by illuminating a sample in a pinpoint manner with a combination of a spotlight source of a laser beam, etc. and a pinhole. Additionally, the pinhole as a spatial filter is arranged on the front side of a photodetector, noise light existing on the same plane as the measurement point is image-formed in the periphery of the pinhole, and light coming from a plane shifted from the measurement point in the direction of an optical axis is broadened with an objective lens before the pinhole, whereby light passing through the pinhole is suppressed. As a result, light beams reflected from and passing through other than the measurement point are cut, so that only one point in three-dimensional space can be measured with a photoelectric converter.
In the meantime, it is also known that the confocal scanning microscope has a high resolution in the direction of the optical axis. That is, the intensity of measured light increases when a focus is achieved on the optical axis, but almost becomes zero when the focus is not achieved. Accordingly, a plurality of sample images sliced on a plane vertical to a Z direction can be obtained by moving a focus with predetermined pitches in the Z direction, which becomes the optical axis direction, while performing two-dimensional scanning (plane scanning) the spotlight on the plane of the sample. These sample images represent a stereoscopic (three-dimensional) form of the sample. Accordingly, the stereoscopic image of this sample can be built from these sliced images (two-dimensional confocal images) thus obtained in respective positions of the sample in the Z direction (hereinafter abbreviated to “respective Z positions”).
There may be cases where the three-dimensional partial region in the stereoscopic image thus built is targeted, and rescanning or a three-dimensional analysis (a calculation of the volume of the target region, or the like) is performed for the target region. At this time, how to determine and extract the three-dimensional target region becomes significant.
As a general method for determining or extracting a three-dimensional target region, there is a method for determining or extracting a target region for each image sliced in each Z position of a sample, and three-dimensionally concatenating determined or extracted regions. If a determination of a specified region is made for a sliced image according to such a method, a person who makes the determination draws a circle, etc. on a display unit such as a monitor, etc., on which a two-dimensional sliced image is displayed, by operating an input device such as a mouse, etc. of a computer, and the region indicated by the drawing is recognized as a specified region. The determination of a specified region with such drawing is individually made for a sliced image of a sample in each position, and all of target regions to which the determination is made in sliced images in respective Z positions are lastly concatenated, whereby a determination of a three-dimensional target region is made.
Besides, for example, a technique disclosed by Japanese publication of examined patent applications No. HEI 7-7444 exists as a technique extracting a three-dimensional target region.
This technique is characterized in comprising: a two-dimensional image inputting unit partitioning a three-dimensional image representing a three-dimensional object into a plurality of two-dimensional binary images, and inputting the images; a two-dimensional image labeling processing unit extracting a two-dimensional concatenation component for each of the two-dimensional binary images input by the two-dimensional image inputting unit, and executing a labeling process; a two-dimensional image representative point extracting unit extracting a representative point for each labeling region obtained by the two-dimensional image labeling processing unit; a representative point concatenation detecting unit detecting a concatenation of two-dimensional images whose representative points extracted by the two-dimensional image representative point extracting unit are adjacent, and extracting a concatenation component of a representative point; and a three-dimensional image labeling processing unit executing a labeling process for a three-dimensional concatenation component including the concatenation component of the representative point extracted by the representative point concatenation detecting unit.
With its extraction method, a plurality of two-dimensional images are first obtained by slicing a three-dimensional object with the two-dimensional image inputting unit, and a concatenation component of each two-dimensional image is then obtained with the labeling process. After the two-dimensional concatenation component is obtained, the representative point extracting unit obtains a representative point of each two-dimensional concatenation component. After a concatenation of the obtained representative points in two-dimensional images is detected, all of two-dimensional concatenation components including the representative points are extracted as three-dimensional concatenation components. Then, only a three-dimensional concatenation component of a target region is extracted from among the three-dimensional concatenation components thus obtained.
Besides, as a technique associated with the present invention, for example, there is a technique disclosed by Japanese publication of unexamined patent applications No. HEI 6-94595. With this technique, three-dimensional gray-scale image data of particles is obtained by acquiring gray-scale image data of a tomographic image group, the mutual position relationship of which is clear, of the inside of a three-dimensional object to be examined where the particles are distributed, and the three-dimensional binary image data of the particles is obtained by binarizing the gray-scale level of the data of each pixel. Then, binary image data of each particle is eroded to the binary image data of its central point by three-dimensionally executing a erosion process for the binary image data, and the position of each particle is three-dimensionally determined with this binary image data.